Background-Eosinophils are likely key cells involved in the pathogenesis of asthma and allergic diseases; however, the mechanisms that regulate eosinophil dynamics and functions in mucosal tissues are incompletely understood. IL-33, which is produced by mucosal cells, is a new member of the IL-1 cytokine family. Mice injected with IL-33 display profound mucosal eosinophilia with associated pathologic changes. Although mast cells and Th2 cells express the IL-33 receptor, ST2, the roles of IL-33 and ST2 in eosinophil biology are unknown.

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Summary Scratching triggers skin flares in atopic dermatitis (AD). We demonstrate that scratching of human skin, and tape stripping of mouse skin, causes neutrophil influx. This influx in mice was largely dependent on the generation of leukotriene B4 (LTB4) by neutrophils and their expression of the LTB4 receptor BLT1. Allergic skin inflammation in response to epicutaneous (EC) application of ovalbumin to tape-stripped skin was severely impaired in Ltb4r1−/− mice, and required expression of BLT1 on both T cells and non-T cells. Co-transfer of WT neutrophils, but not neutrophils deficient in BLT1 or the LTB4 synthesizing enzyme LTA4H, restored the ability of WT CD4+ effector T cells to transfer allergic skin inflammation to Ltb4r1−/− recipients. Pharmacologic blockade of LTB4 synthesis inhibited allergic skin inflammation elicited by cutaneous antigen challenge in previously EC-sensitized mice. Our results demonstrate that a neutrophil-T cell axis reliant on LTB4-BLT1 interaction is required for allergic skin inflammation.

Background Cutaneous exposure to food allergens predisposes to food allergy, which is commonly associated with atopic dermatitis (AD). The levels of the epithelial cytokine IL-33 are increased in skin lesions and serum of AD patients. Mast cells (MC) play a critical role in food anaphylaxis and express the IL-33 receptor ST2. The role of IL-33 in MC-dependent food anaphylaxis is unknown. Objective To determine the role and mechanism of action of IL-33 in food anaphylaxis in a model of IgE-dependent food anaphylaxis elicited by oral challenge of epicutaneously (EC)-sensitized mice. Methods WT, ST2-deficient and MC-deficient KitW-sh/W-sh mice were EC sensitized with ovalbumin (OVA) then challenged orally with OVA. Body temperature was measured by telemetry, Il33 mRNA by qPCR, and IL-33, OVA-specific IgE and mouse mast cell protease 1 (mMCP-1) by ELISA. Bone marrow-derived MCs (BMMCs) degranulation was assessed by flow cytometry. Results Il33 mRNA expression was upregulated in tape-stripped mouse skin and scratched human skin. Tape stripping caused local and systemic IL-33 release in mice. ST2 deficiency, as well as ST2 blockade prior to oral challenge, significantly reduced the severity of oral anaphylaxis without affecting the systemic Th2 response to the allergen. Oral anaphylaxis was abrogated in KitW-sh/W-sh mice, and restored by reconstitution with WT, but not ST2-deficient, BMMCs. IL-33 significantly enhanced IgE-mediated degranulation of BMMCs in vitro. Conclusion IL-33 is released following mechanical skin injury, enhances IgE-mediated MC degranulation, and promotes oral anaphylaxis following EC sensitization by targeting MCs. IL-33 neutralization may be useful in treating food anaphylaxis in AD patients.

Here we report new photoactivable antibody binding proteins, which site-selectively capture antibodies and form covalent conjugates with captured antibodies upon irradiation. The proteins allow the site-selective tagging and/or immobilization of antibodies with a highly preferred orientation and omit the need for prior antibody modifications. The minimal Fc-binding domain of protein G, a widely used antibody binding protein, was genetically and chemically engineered to contain a site-specific photo cross-linker, benzophenone. In addition, the domain was further mutated to have an enhanced Fc-targeting ability. This small engineered protein was successfully cross-linked only to the Fc region of the antibody without any nonspecific reactivity. SPR analysis indicated that antibodies can be site-selectively biotinylated through the present photoactivable protein. Furthermore, the system enabled light-induced covalent immobilization of antibodies directly on various solid surfaces, such as those of glass slides, gold chips, and small particles. Antibody coupling via photoactivable antibody binding proteins overcomes several limitations of conventional approaches, such as random chemical reactions or reversible protein binding, and offers a versatile tool for the field of immunosensors.

Background-Atopic dermatitis (AD) is characterized by local and systemic Th2 responses to cutaneously introduced allergens and is a risk factor for asthma. Blockade of Th2 cytokines has been suggested as therapy for AD.

Eosinophils produce and release various proinflammatory mediators and also show immunomodulatory and tissue remodeling functions; thus, eosinophils may be involved in the pathophysiology of asthma and other eosinophilic disorders as well as host defense. Several major questions still remain. For example, how do human eosinophils become activated in diseased tissues or at the site of an immune response? What types of host immunity might potentially involve eosinophils? Herein, we found that human eosinophils react vigorously to a common environmental fungus, Alternaria alternata, which is implicated in the development and/or exacerbation of human asthma. Eosinophils release their cytotoxic granule proteins, such as eosinophil-derived neurotoxin and major basic protein, into the extracellular milieu and onto the surface of fungal organisms and kill the fungus in a contact-dependent manner. Eosinophils use their versatile β2 integrin molecule, CD11b, to adhere to a major cell wall component, β-glucan, but eosinophils do not express other common fungal receptors, such as dectin-1 and lactosylceramide. The I-domain of CD11b is distinctively involved in the eosinophils’ interaction with β-glucan. Eosinophils do not react with another fungal cell wall component, chitin. Because human eosinophils respond to and kill certain fungal organisms, our findings identify a previously unrecognized innate immune function for eosinophils. This immune response by eosinophils may benefit the host, but, in turn, it may also play a role in the development and/or exacerbation of eosinophil-related allergic human diseases, such as asthma.

Eosinophils and their products are likely important in the pathophysiology of allergic diseases, such as bronchial asthma, and in host immunity to parasitic organisms. However, the mechanisms for proinflammatory mediator release by eosinophils are poorly understood. CD66b (CEACAM8, CGM6, NCA-95) is a single chain, GPI-anchored, highly glycosylated protein belonging to the carcinoembryonic Ag supergene family. CD66b is an activation marker for human granulocytes; however, its biological functions are largely unknown in eosinophils. We found that CD66b is highly expressed on the surface of human peripheral blood eosinophils isolated from healthy individuals. Engagement of CD66b, but not CD66a, by mAb or a natural ligand, galectin-3, activated a Src kinase family molecule, hemopoietic cell kinase (Hck), and induced cellular adhesion, superoxide production, and degranulation of eosinophils. CD66b molecules were localized in lipid rafts, and disruption of lipid rafts or removal of the GPI anchor inhibited the adhesion and activation of eosinophils. Importantly, CD66b was constitutively and physically associated with a β2 integrin, CD11b, and cross-linking of CD66b induced a striking clustering of CD11b molecules. Thus, CD66b molecules are involved in regulating adhesion and activation of eosinophils, possibly through their localization in lipid rafts and interaction with other cell surface molecules, such as CD11b. Binding of exogenous or endogenous carbohydrate ligands(s) to CD66b may be important in the release of proinflammatory mediators by human eosinophils.